梅花鹿FSHβ亚基基因克隆与融合表达

Gene Cloning and Fusion Expression of Cervus nippon FSHβ-subunit in E. coli

以原构建的克隆载体为模板,PCR扩增梅花鹿FSHβ亚基基因,TA克隆后经双酶切插入表达载体pGEX-6P-2,阳性克隆导入E.coliBL21,IPTG诱导表达GST-FSHβ融合蛋白,SDS-PAGE进行分析鉴定。结果显示,FSHβPCR产物大小约410 bp,测序结果与GenBank序列一致,成功构建了重组表达载体pGEX-6P-2-FSHβ,融合蛋白经SDS-PAGE分析,在相对分子量39.5 kD处,出现特异性蛋白条带,说明梅花鹿FSHβ亚基基因片段已在E.coliBL21中成功表达了FSHβ-GST融合蛋白,融合蛋白功能有待进一步分析。

FSHβ-subunit gene segments were cloned by PCR, from the original constructed vector. Then the PCR product was TA cloned, sequenced and inserted into the carrier pGEX-6P-2. The recombinant plasmids were identified by restricted enzymes digestion, and then the positive clones were transformed into E. coli BL21. GST-FSHβ fusion proteins were expressed via the induction of IPTG, and detected by SDS-PAGE. Results showed that the PCR product was about 410 bp in size and sequencing result was identical with GenBank＇s report. The pGEX-6P-2-FSHβ positive clone produced a 39.5 kD fusion protein on SDS-PAGE gel, and the function of which needs further study.